Solvent extraction of recombinant interferon alpha-2b from inclusion bodies and efficient refolding at high protein concentrations.

Interferon alpha-2b (IFNα-2b) is an essential cytokine widely used in hepatitis and cancer treatment. This paper presents a novel protocol for purification and efficient refolding of recombinant interferon alpha-2b (IFNα-2b) expressed as insoluble inclusion bodies in Escherichia coli. Purification of IFNα-2b from solubilized inclusion bodies was performed by solvent extraction using 2-butanol. Refolding conditions were optimized using the response surface method (RSM). Under optimized conditions, refolding yield of solvent-extracted IFNα-2b was 1.5 fold higher than refolding yield of unpurified IFNα-2b. High-concentration protein refolding was carried out by pulse-fed method, and refolding yield of 75% was achieved at a protein concentration of 300 µg ml-1. Under optimized conditions, 259.16 mg of purified IFNα-2b with the biological activity of 2.4 × 108 IU mg-1 was achieved per liter of bacterial culture. The developed protocol provides an efficient production process of high-quality IFNα-2b suitable for research and pharmaceutical applications.

Promising insights into the kosmotropic effect of magnetic nanoparticles on proteins: The pivotal role of protein corona formation.

Increasing concerns about biosafety of nanoparticles (NPs) has raised the need for detailed knowledge of NP interactions with biological molecules especially proteins. Herein, the concentration-dependent effect of magnetic NPs (MNPs) on bovine serum albumin and hen egg white lysozyme was explored. The X-ray diffraction patterns, zeta potential, and dynamic light scattering measurements together with scanning electron microscopy images were employed to characterize MNPs synthesized through coprecipitation method. Then, we studied the behavior of two model proteins with different surface charges and structural properties on interaction with Fe3 O4 . A thorough investigation of protein-MNP interaction by the help of intrinsic fluorescence at different experimental conditions revealed that affinity of proteins for MNPs is strongly affected by the similarity of protein and MNP surface charges. MNPs exerted structure-making kosmotropic effect on both proteins under a concentration threshold; however, binding strength was found to determine the extent of stabilizing effect as well as magnitude of the concentration threshold. Circular dichroism spectra showed that proteins with less resistance to conformational deformations are more prone to secondary structure changes upon adsorption on MNPs. By screening thermal aggregation of proteins in the presence of Fe3 O4 , it was also found that like chemical stability, thermal stability is influenced to a higher extent in more strongly bound proteins. Overall, this report not only provides an integrated picture of protein-MNP interaction but also sheds light on the molecular mechanism underling this process.

Production of Vitamin D3 Enriched Biomass of Saccharomyces Cerevisiae as A Potential Food Supplement: Evaluation and Optimization of Culture Conditions Using Plackett-Burman and Response Surface Methodological Approaches.

Vitamin D deficiency causes osteoporosis, osteopenia, fractures, rickets, and more recently is linked with some chronic illnesses such as cancer. Because of the safety and probiotic properties of the yeast Saccharomyces cerevisiae, we hypothesized that yeast cells enriched with cholecalciferol (vitamin D3) could represent a solution for prevention or treatment of vitamin D deficiency. In this study S. cerevisiae was used as a vitamin D3 accumulator for the first time and the optimal conditions for enrichment of S. cerevisiae were determined. The Plackett-Burman screening studies were used for selection of the most important factors affecting cholecalciferol entrapment. Response surface methodology was employed for optimization of cholecalciferol accumulation in S. cerevisiae cells by using Box-Behnken design. A modified quadratic polynomial model fit the data appropriately. The optimal points of variables to maximize the response were cholecalciferol initial concentration of 358021.16 IU/mL, tryptone concentration of 1.82 g/L, sucrose concentration of 7.13 % (w/v), and shaking speed of 140.46 rpm. The maximum amount of cholecalciferol in dry cell weight of S. cerevisiae was 4428.11 IU/g. The cholecalciferol entrapment in yeast biomass increased about two-folds in optimized condition which indicates efficiency of optimization.